In response to injury, cancer, microbial invasion, and the like, humans and other animals mount inflammatory responses to control the pathological condition and to initiate a repair process. During inflammation, various immune cells including T-lymphocytes, neutrophils and macrophages are recruited to the site where they produce cytokines to facilitate the immune response. Among these cytokines, tumor necrosis factor-α (TNF-α) is one of the major proinflammatory proteins that mediates the immune response. Although the effects of proinflammatory cytokines are protective, their overproduction may have adverse effects to the host. In fact, uncontrolled induction of proinflammatory cytokines can lead to complications such as hypotension, organ failure and even death1,2.
During an acute phase of infection such as in the case of sepsis, uncontrolled production of TNF-α is well known to cause deleterious effects to the host. Sepsis is the second most common cause of death in non-coronary intensive care units and the tenth leading cause of death overall in high-income countries3. The clinical outcome of infection leading to sepsis is primarily associated with the excessive stimulation of the host immune cells, particularly monocytes or macrophages, by bacterial endotoxins (e.g., lipopolysaccharide [LPS])4, 5, 6. Macrophages overstimulated by LPS also produce high levels of mediators such as interleukin-1 (IL-1), IL-6, and TNF-α7. These mediators are implicated in the pathogenesis of sepsis and found to be contributing factors to the demise of the host.
In addition to its role in acute phase response, TNF-α has been shown to be involved in the progression of various chronic diseases including tumorigenesis and rheumatoid arthritis (RA). The dysregulation of TNF-α production has been demonstrated to be involved in different stages of tumorigenesis including initiation of tumor growth8, cell proliferation9 and invasion10. For tumor cell proliferation, TNF-α upregulates specific growth factors to mediate the malignant growth. The cytokine promotes angiogenesis that supports tumor migration, and thus plays a key role in tumor metastasis. For example, glioblastoma migration and induction of matrix metalloproteinases (MMP) are significantly enhanced in response to TNF-α effects11. This induction of MMP in glioblastoma T98G cells can be reversed by treatment of the cells with interferon-g12.
The uncontrolled production of TNF-α is associated with many acute and chronic neurodegenerative conditions, including stroke, brain trauma, spinal cord injury, amyotrophic lateral sclerosis (ALS), Huntington's disease, Alzheimer's disease, and Parkinson's disease. Studies show that TNF-α rapidly upregulates in the brain after injury, and it plays a pivotal role in inflammatory processes. For instance, studies show that cerebrovascular diseases including ischemic stroke are associated with inflammation mediated responses in the neural cells. Also, the enhanced expression of TNF-α has been found in association with glial cells in the substantia nigra of patients with Parkinson's disease.
The toxic effects of TNF-α and its role as a mediator of focal ischemia may involve many other mechanisms in addition to inflammation. For example, increased TNF-α in the brain and blood in response to lipopolysaccharide (LPS) appears to contribute to increased brain stem thrombosis and hemorrhage, and increased stroke sensitivity/risk. Additionally, TNF-α increases blood-brain barrier permeability and produces pial artery constriction, which can contribute to focal ischemic brain injury. Further, there appears to be a direct toxic effect of TNF-α on capillaries. Specifically, TNF-α increases capillary permeability and opens the blood-brain barrier, apparently by increasing matrix-damaging metalloproteinase (gelatinase B) production, which is also expressed early after focal stroke. TNF-α also causes damage to myelin and oligodendrocytes and increases astrocytic proliferation, thus potentially contributing to demyelination and reactive gliosis during brain injury.
Further examples of acute and chronic disease pathogenesis mediated by TNF-α include rheumatoid arthritis and inflammatory bowel diseases. Patients with rheumatoid arthritis have a low grade insidious inflammation in the synovial tissues. It is known that overproduction of TNF-α at the inflamed joint leads to slow destruction of the joint cartilage and surrounding bone.
Additionally, inflammatory responses including TNF-α production may play an important role in the pathogenesis of cerebrovascular diseases including ischemic stroke and cardiovascular diseases (CVD). It has been suggested that TNF-α may destabilize atherogenesis and atherosclerotic plaques leading to their rupture, resulting in myocardial infarction or stroke in CVD patients.
Furthermore, studies have shown that disease pathogenesis mediated by TNF-α can be associated with microbial, bacterial and viral infections. Cytokines such as TNF-α play a role in defending against the invading pathogens such as, for example, mycobacteria, influenza viruses, SARS-coronavirus and retroviruses including HIV. However, many microbes and viruses have also developed various immunosuppressive mechanisms that cause dysfunction of protein signaling kinases and transcription factors as well as other components involved in the TNF-α signaling pathway13, 14, 15, 16, 17, 18.
In addition to uncontrolled production of TNF-α, immune cells in a diseased condition are activated by cytokines, microbial compounds or both to generate nitric oxide (NO). Generation of nitric oxide is a feature of genuine immune-system cells such as dendritic cells, NK cells, mast cells and phagocytic cells including monocytes, macrophages, microglia, Kupffer cells, eosinophils, and neutrophils as well as other cells involved in immune reactions. Many targets of NO are themselves regulatory molecules, for example transcription factors and components of various signaling cascade.
Additional keys mediators involved in immune response further include interferons such as Interferon-gamma (IFN-γ); the interleukin family such as Interleukin-1 (IL-1), Interleukin-2 (IL-2), Interleukin-3 (IL-3), Interleukin-4 (IL-4), Interleukin-5 (IL-5), Interleukin-6 (IL-6), Interleukin-7 (IL-7), Interleukin-8 (IL-8), Interleukin-9 (IL-9), Interleukin-10 (IL-10), Interleukin-11 (IL-11), Interleukin-12 (IL-12), Interleukin-13 (IL-13), Interleukin-14 (IL-14), Interleukin-15 (IL-15), Interleukin-16 (IL-16), Interleukin-17 (IL-17), Interleukin-18 (IL-18), Interleukin-19 (IL-19), Interleukin-20 (IL-20), Interleukin-21 (IL-21), Interleukin-22 (IL-22), Interleukin-23 (IL-23), Interleukin-24 (IL-24), Interleukin-25 (IL-25), Interleukin-26 (IL-26), Interleukin-27 (IL-27), Interleukin-28 (IL-28), Interleukin-29 (IL-29), Interleukin-30 (IL-30), Interleukin-31 (IL-31), Interleukin-32 (IL-32), Interleukin-33 (IL-33), Interleukin-34 (IL-34), Interleukin-35 (IL-35); the interleukin receptor family; the macrophage inflammatory protein family such as macrophage inflammatory protein 2 (MIP-2) and macrophage inflammatory protein 1α (MIP-1α); macrophage colony-stimulating factor (M-CSF); and monocyte chemotactic protein-1 (MCP-1).
Targeting the uncontrolled production of inflammatory mediators such as TNF-α and nitric oxide has played an increasing role in treating inflammatory and immune conditions. Use of exogenous anti-inflammatory therapeutics would be particularly desirable to control the adverse effects of immune over-activation. In recent years, immunotherapeutics have been developed that aim at the neutralization of TNF-α and suppression of its undesirable proinflammatory effects. For example, as TNF-α exacerbates focal ischemic injury in neurodegenerative diseases, agents for blocking endogenous TNF-α have been shown to be neuroprotective. These agents include soluble TNF-α receptor (Enbrel) and anti-TNF-α antibody (Infliximab). Despite their novelty and efficacy in the arrest of disease progression, they are very expensive therapeutic regimens.
In addition, non-steroid anti-inflammatory drugs (NSAIDs) including aspirin, ibuprofen, and indomethacin are well-known in ameliorating acute and chronic pain associated with inflammatory diseases such as rheumatoid arthritis and inflammatory bowel disease. However, they are not effective in the treatment of advanced stages of rheumatoid arthritis and related autoimmune diseases. For those conditions, steroids and cytotoxic drugs such as methotrexate and cyclophosphamide are used. These drugs are associated with severe adverse effects including gastrointestinal irritation, severe bleeding, and bone marrow suppression.
Stroke is the leading cause of adult disability and the third most prevalent cause of mortality worldwide. There is increasing evidence that inflammation accounts for the progression of ischemic stroke by causing neuronal damages19,20, 21. Microglia cells, the resident macrophages in the brain, are activated during ischemia. In response to the compromised state of oxygen supply, the cells generate various inflammatory mediators including TNF-α, nitric oxide, IL-6 and IL-122. Thus, the development of novel therapies directed towards the inhibition of pathological production of TNF-α and NO is needed to aid in the treatment of these acute and chronic immune diseases.